Cloning goat producing human lactoferrin with genetically modified donor cells selected by single or dual markers / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 1482-1491, 2012.
Article
in Chinese
| WPRIM
| ID: wpr-233278
ABSTRACT
We compared the efficiency of cloning goat using human lactoferrin (hLF) with genetically modified donor cells marked by single (Neo(r)) or double (Neo(r)/GFP) markers. Single marker expression vector (pBLC14) or dual markers expression vector (pAPLM) was delivered to goat fetal fibroblasts (GFF), and then the transgenic GFF was used as donor cells to produce transgenic goats. Respectively, 58.8% (20/34) and 86.7% (26/30) resistant cell lines confirmed the transgenic integration by PCR. Moreover, pAPLM cells lines were subcultured with several passages, only 20% (6/30) cell lines was observed fluorescence from each cell during the cell passage. Somatic cell nuclear transfer using the donor cells harbouring pBLC14 or pAPLM construct, resulting in a total of 806 reconstructed embryos, a pregnancy rate at 35 d (53.8%, 39.1%) and 60 d (26.9%, 21.7%), and an offspring birth rate (1.9%, 1.4%) with 5 and 7 newborn cloned goats, respectively. Transgene was confirmed by PCR and southern-blot in all cloned offspring. There were no significant differences at the reconstructed embryo fusion rates, pregnancy rates and the birth rate (P > 0.05) between single and double markers groups. The Neo(r)/GFP double markers could improve the reliability for accurately and efficiently selecting the genetically modified donor cells. No adverse effect was observed on the efficiency of transgenic goat production by SCNT using somatic cells transfected with double (Neo(r)/GFP) markers vector.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Recombinant Proteins
/
Goats
/
Animals, Genetically Modified
/
Neomycin
/
Transfection
/
Genetic Markers
/
Embryology
/
Cloning, Molecular
/
Cloning, Organism
/
Cell Biology
Limits:
Animals
/
Humans
Language:
Chinese
Journal:
Chinese Journal of Biotechnology
Year:
2012
Type:
Article
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